Normal telomere length and chromosomal end capping in poly(ADP-ribose) polymerase–deficient mice and primary cells despite increased chromosomal instability
TL;DR: The results presented here indicate that PARp-1 does not play a major role in regulating telomere length or in telomeric end capping, and the chromosomal instability of PARP-1−/− primary cells can be explained by the repair defect associated to PARP -1 deficiency.
Abstract: Poly(ADP-ribose) polymerase (PARP)-1, a detector of single-strand breaks, plays a key role in the cellular response to DNA damage. PARP-1-deficient mice are hypersensitive to genotoxic agents and display genomic instability due to a DNA repair defect in the base excision repair pathway. A previous report suggested that PARP-1-deficient mice also had a severe telomeric dysfunction consisting of telomere shortening and increased end-to-end fusions (d'Adda di Fagagna, F., M.P. Hande, W.-M. Tong, P.M. Lansdorp, Z.-Q. Wang, and S.P. Jackson. 1999. NAT: Genet. 23:76-80). In contrast to that, and using a panoply of techniques, including quantitative telomeric (Q)-FISH, we did not find significant differences in telomere length between wild-type and PARP-1(-/)- littermate mice or PARP-1(-/)- primary cells. Similarly, there were no differences in the length of the G-strand overhang. Q-FISH and spectral karyotyping analyses of primary PARP-1(-/)- cells showed a frequency of 2 end-to-end fusions per 100 metaphases, much lower than that described previously (d'Adda di Fagagna et al., 1999). This low frequency of end-to-end fusions in PARP-1(-/)- primary cells is accordant with the absence of severe proliferative defects in PARP-1(-/)- mice. The results presented here indicate that PARP-1 does not play a major role in regulating telomere length or in telomeric end capping, and the chromosomal instability of PARP-1(-/)- primary cells can be explained by the repair defect associated to PARP-1 deficiency. Finally, no interaction between PARP-1 and the telomerase reverse transcriptase subunit, Tert, was found using the two-hybrid assay.
Citations
More filters
TL;DR: Findings argue that there remains a possibility that PARPs play a role in Arabidopsis telomere biology, but that the contribution is a minor one.
Abstract: Maintaining the length of the telomere tract at chromosome ends is a complex process vital to normal cell division. Telomere length is controlled through the action of telomerase as well as a cadre of telomere-associated proteins that facilitate replication of the chromosome end and protect it from eliciting a DNA damage response. In vertebrates, multiple poly(ADP-ribose) polymerases (PARPs) have been implicated in the regulation of telomere length, telomerase activity and chromosome end protection. Here we investigate the role of PARPs in plant telomere biology. We analyzed Arabidopsis thaliana mutants null for PARP1 and PARP2 as well as plants treated with the PARP competitive inhibitor 3-AB. Plants deficient in PARP were hypersensitive to genotoxic stress, and expression of PARP1 and PARP2 mRNA was elevated in response to MMS or zeocin treatment or by the loss of telomerase. Additionally, PARP1 mRNA was induced in parp2 mutants, and conversely, PARP2 mRNA was induced in parp1 mutants. PARP3 mRNA, by contrast, was elevated in both parp1 and parp2 mutants, but not in seedlings treated with 3-AB or zeocin. PARP mutants and 3-AB treated plants displayed robust telomerase activity, no significant changes in telomere length, and no end-to-end chromosome fusions. Although there remains a possibility that PARPs play a role in Arabidopsis telomere biology, these findings argue that the contribution is a minor one.
24 citations
Cites background from "Normal telomere length and chromoso..."
...Furthermore, mice deficient in Tankyrase1 or Tankyrase2 have normal telomeres over multiple generations [35,98,99], and while some investigators report telomere shortening and other telomere aberrations in PARP-deficient mice [56,91,93], others have been unable to detect telomere defects [55,92,100]....
[...]
TL;DR: The level of PARP-1 gene dosage correlates with PARP activity and the in vivo level of poly(ADP-ribosyl)ation, which could explain the mechanism by which PARp-1 haploinsufficiency affects centrosome duplication and chromosome stability.
22 citations
TL;DR: Results indicate that PARP-1 and p53 cooperate in the suppression of tumorigenesis by maintaining genomic integrity after DNA damage through the activation of a G1/S cell cycle checkpoint the initiation of DNA repair and the induction of cell death.
Abstract: Cells that lack PARP-1 activity are limited in their ability to repair DNA single strand breaks and respond to DNA damage with a strong accumulation of p53 and enhanced rates of apoptotic cell death. We have generated combinatorial mutant mice that both lack p53 and PARP-1 activity due to the expression of a dominant negative PARP-1 allele targeted to T-cells by the lck promoter. Here we report that these double mutant mice develop T-cell lymphoma at a significantly reduced latency period compared to single p53 null mice that are already cancer prone. We demonstrate that the absence of p53 does not only protect T-cells from lck-PARP-DBD transgenic mice from apoptosis but also abrogates the DNA damage induced cell cycle arrest in the G1 phase. T-cells from double mutant mice continue to proliferate after the induction of DNA strand breaks, are limited in their DNA repair capacity and cannot be eliminated by apoptosis. These results indicate that PARP-1 and p53 cooperate in the suppression of tumorigenesis by maintaining genomic integrity after DNA damage through the activation of a G1/S cell cycle checkpoint the initiation of DNA repair and the induction of cell death.
22 citations
Cites background from "Normal telomere length and chromoso..."
...…DBD transgenics show severe defects in single strand break repair and elevated sister chromatid exchange (Beneke et al., 2000; Conde et al., 2001; Samper et al., 2001) the development of spontaneous tumors has never been observed in these mouse mutants (data shown here and Wang et al., 1995;…...
[...]
TL;DR: PARP-1 modulates the telomerase activity by altering poly(ADP-ribosyl)ation of TERT and/or the expression of TEP1/TP1 and such modification was decreased in cells with reduced PARp-1 expression.
Abstract: Inhibitors of poly(ADP-ribose) polymerase (PARP) have been shown to reduce the telomerase activity in cultured cells. To find out the specific member of the PARP family, which participates in the regulation of telomerase activity, in the present investigation, we knocked down the PARP-1 gene by siRNA in HeLa cells and studied the telomerase activity. Reduction of expression of PARP-1 by siRNA increased cellular NAD + level and decreased general poly(ADP-ribosyl)ation of proteins. Telomerase activity decreased in cells with knocked down PARP-1 gene. Besides, we observed that telomerase reverse transcriptase (hTERT) was poly(ADP-ribosyl)ated in HeLa cells and such modification was decreased in cells with reduced PARP-1 expression. In addition, the expression of telomerase-associated protein 1 (TEP1/TP1) subunit of human telomerase holoenzyme reduced significantly in PARP-1 knock down HeLa cells. Thus, PARP-1 modulates the telomerase activity by altering poly(ADP-ribosyl)ation of TERT and/or the expression of TEP1/TP1.
22 citations
TL;DR: A novel function and mechanism underlying cytoplasmic PARP‐1, distinct from nuclear PARP•1, in regulating DR5‐activated apoptosis is identified and supported by an innovative application of available PARP inhibitors or new cytop lasmic parp‐1 antagonists to enhance TRAIL therapy for TRAIL‐resistant pancreatic cancers.
Abstract: The poly(ADP-ribose) polymerases (PARP) play important roles in repairing damaged DNA during intrinsic cell death. We recently linked PARP-1 to death receptor (DR)-activated extrinsic apoptosis, the present studies sought to elucidate the function of cytoplasmic PARP-1 in pancreatic cancer tumorigenesis and therapy. Using human normal and pancreatic cancer tissues, we analyzed the prevalence of cytoplasmic PARP-1 expression. In normal human pancreatic tissues, PARP-1 expression was present in the nucleus; however, cytoplasmic PARP-1 expression was identified in pancreatic cancers. Therefore, cytoplasmic PARP-1 mutants were generated by site-direct mutagenesis, to determine a causative effect of cytoplasmic PARP-1 on pancreatic cancer tumorigenesis and sensitivity to therapy with TRA-8, a humanized DR5 antibody. PARP-1 cytoplasmic mutants rendered TRA-8 sensitive pancreatic cancer cells, BxPc-3 and MiaPaCa-2, more resistant to TRA-8-induced apoptosis; whereas wild-type PARP-1, localizing mainly in the nucleus, had no effects. Additionally, cytoplasmic PARP-1, but not wild-type PARP-1, increased resistance of BxPc-3 cells to TRA-8 therapy in a mouse xenograft model in vivo. Inhibition of PARP enzymatic activity attenuated cytoplasmic PARP-1-mediated TRA-8 resistance. Furthermore, increased cytoplasmic PARP-1, but not wild-type PARP-1, was recruited into the TRA-8-activated death-inducing signaling complex and associated with increased and sustained activation of Src-mediated survival signals. In contrast, PARP-1 knockdown inhibited Src activation. Taken together, we have identified a novel function and mechanism underlying cytoplasmic PARP-1, distinct from nuclear PARP-1, in regulating DR5-activated apoptosis. Our studies support an innovative application of available PARP inhibitors or new cytoplasmic PARP-1 antagonists to enhance TRAIL therapy for TRAIL-resistant pancreatic cancers.
21 citations
References
More filters
TL;DR: P16 seems to act in a regulatory feedback circuit with CDK4, D-type cyclins and retinoblastoma protein, and inhibits the catalytic activity of theCDK4/cyclin D enzymes.
Abstract: The division cycle of eukaryotic cells is regulated by a family of protein kinases known as the cyclin-dependent kinases (CDKs). The sequential activation of individual members of this family and their consequent phosphorylation of critical substrates promotes orderly progression through the cell cycle. The complexes formed by CDK4 and the D-type cyclins have been strongly implicated in the control of cell proliferation during the G1 phase. CDK4 exists, in part, as a multi-protein complex with a D-type cyclin, proliferating cell nuclear antigen and a protein, p21 (refs 7-9). CDK4 associates separately with a protein of M(r) 16K, particularly in cells lacking a functional retinoblastoma protein. Here we report the isolation of a human p16 complementary DNA and demonstrate that p16 binds to CDK4 and inhibits the catalytic activity of the CDK4/cyclin D enzymes. p16 seems to act in a regulatory feedback circuit with CDK4, D-type cyclins and retinoblastoma protein.
3,716 citations
"Normal telomere length and chromoso..." refers methods in this paper
...As control for the two-hybrid assay we show interaction between the cell cycle proteins CDK4 and p16 as described previously (Table VI; Serrano et al., 1993)....
[...]
TL;DR: The DNA of telomeres—the terminal DNA-protein complexes of chromosomes—differs notably from other DNA sequences in both structure and function, and has been shown to be essential for telomere maintenance and long-term viability.
Abstract: The DNA of telomeres--the terminal DNA-protein complexes of chromosomes--differs notably from other DNA sequences in both structure and function. Recent work has highlighted its remarkable mode of synthesis by the ribonucleoprotein reverse transcriptase, telomerase, as well as its ability to form unusual structures in vitro. Moreover, telomere synthesis by telomerase has been shown to be essential for telomere maintenance and long-term viability.
3,139 citations
TL;DR: Electron microscopy reported here demonstrated that TRF2 can remodel linear telomeric DNA into large duplex loops (t loops) in vitro, which may provide a general mechanism for the protection and replication of telomeres.
2,413 citations
TL;DR: Results indicate that telomerase is essential for telomere length maintenance but is not required for establishment of cell lines, oncogenic transformation, or tumor formation in mice.
2,066 citations
"Normal telomere length and chromoso..." refers background or methods in this paper
...These Robertsonian fusions are different from those found in late generation telomerase-deficient mice that have critically short telomeres (Blasco et al., 1997)....
[...]
...5 embryos derived from heterozygous crosses as described (Blasco et al., 1997)....
[...]
...Telomeres have an essential role in chromosome stability and are proposed to be biological determinants in the processes of tumorigenesis and aging (for reviews see Blackburn, 1991; Autexier and Greider, 1996; Greider, 1996; Blasco et al., 1997; Lee et al., 1998)....
[...]
...Then the plugs were digested with MboI overnight and run in a pulse field gel electrophoresis as described (Blasco et al., 1997)....
[...]
TL;DR: The history and present situation of Spanish language, culture, literature, cuisine, tourism, and more are explored in more detail in this booklet.
Abstract: TELOMERES DEFINED . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 579 TELOMERE FUNCTION . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 580 SEQUENCE AND STRUCTURE OF TELOMERES . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 581 SOLUTIONS FOR REPLICATION OF DNA TERMINI . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 586 STRUCTURE OF SUBTELOMERIC REGIONS.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 589 FORMA TION OF TELOMERES . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ... . . . . . .. 591 PROTEINS THAT INTERACT WITH TELOMERES . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 594 ARE TELOMERES REALLY ESSENTIAL? . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 597 FUTURE PROSPECTS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 598
1,923 citations
"Normal telomere length and chromoso..." refers background in this paper
...Vertebrate telomeres consist of tandem repeats of the sequence TTAGGG (for review see Blackburn, 1991)....
[...]
...Telomeres have an essential role in chromosome stability and are proposed to be biological determinants in the processes of tumorigenesis and aging (for reviews see Blackburn, 1991; Autexier and Greider, 1996; Greider, 1996; Blasco et al., 1997; Lee et al., 1998)....
[...]